Fig. 4.

The predicated functions of isolated nucleic acid ligand. (A) Effect of simultaneous treatment with aptamer and UTP on wild-type P2RY2 activity. The dose–response curve using UTP is shown (Left). Cells overexpressing wild-type P2RY2 were exposed to the mixture of aptamer c37_8-40 and UTP at the indicated concentrations (Right). The values are expressed as RLU (in %) to the 2.5 μM (Left) and the 100 nM (Right) UTP level without aptamer. Data represent the mean ± SD (n = 3 independent experiments). (B) Effect of Gαq inhibitor on aptamer-induced calcium signaling. The effect of YM-254890 on the calcium signaling stimulated by 10 nM UTP was shown as RLU (in %) to the 10 nM UTP level without inhibitor (Left). Likewise, inhibitory effect of YM-254890 at 100 nM, which is sufficient to block calcium signaling activated by 10 nM UTP, was examined for the calcium signaling stimulated by 2.5 μM aptamers. The values are expressed as RLU (in %) to the 10 nM UTP level without aptamer. Control group is not exposed to the aptamers and inhibitor. Data represent the mean ± SD (n = 3 independent experiments). ^#P < 0.05 versus control group; and *P < 0.05 between indicated groups. (C) Schematic drawing of the predicted functions of the aptamer. Our results suggest that c37_8-40 aptamer binds to an allosteric site of P2RY2 and exhibits dual function, agonist, and PAM, depending on whether the ligand UTP is prebound to the receptor.